Several converging lines of inquiry suggest that delayed neuronal loss associated with diseases such as stroke or spinal cord trauma occurs by a process known as apoptosis. Enhanced understanding of the common and distinct mechanisms by which apoptosis is triggered may thus reveal novel therapeutic approaches to limit neurologic disability associated with these diseases. We previously demonstrated that apoptotic death triggered by oxidative stress in cortical neurons can be abrogated by the antioxidant, N-acetylcysteine, NAC, a drug commonly used in humans to treat acetaminophen toxicity. To determine whether NAC is broadly applicable as an anti-apoptotic agent, we examined the effects of this antioxidant in another paradigm of neuronal apoptosis: infection with Sindbis Virus (SV). SV appears to be a good model to study general mechanisms of neuronal apoptosis as many of the agents which prevent SV-induced death (e.g. bcl-2, proteosome inhibitors) prevent growth factor deprivation-induced apoptosis in sympathetic neurons. We found that NAC prevented SV-induced apoptosis, but that much higher concentrations of the drug were required (30 mM). In preliminary studies, we correlated the protective effects of NAC (in two cultured cell lines) with its ability to inhibit SV-induced translocation of the redox-sensitive transcription factor, NF-kappa B (NFkB). Inhibition of NF-kB DNA binding activity directly using molecular approaches prevented SV-induced death in one or two cell lines examined. Subsequent studies revealed that SV-induces NF-kB and apoptosis in primary neurons. These studies raise two specific hypothesis: Hypothesis #1: NFkB is required for cell death in cortical neuronal cultures; and Hypothesis #2: SV induces NFkB and apoptosis in cortical neuronal cultures through an oxidant second messenger (e.g., hydrogen peroxide). In Specific Aim #1, we will investigate whether SV-induced activation of NFkB is required for apoptosis in primary neuronal cultures. We will utilize phosphorothioate double stranded oligonucleotides containing NFkB binding sites as decoys to inhibit binding of NFkB to authentic DNA sites. Additionally, we will utilize SV as a vector not only to initiate apoptosis, but also to deliver a protein inhibitor of NFkB activation. In Specific Aim 2, we will investigate whether SV-induced NFkB activation or apoptosis involves the generation of oxidant second messenger. These studies promise to enhance our understanding of apoptotic mechanisms in neurons and give insights into novel approaches to abrogate apoptosis in the nervous system.